The hematopoietic stem cell (HSC) compartment is heterogeneous and includes lymphoid biased (Ly-HSCs) and myeloid biased (My-HSCs) subpopulations. It has been assumed that the number of Ly-HSCs declines with age and that this explains why lymphopoiesis is attenuated in the elderly. However, this view is based on assessing stem cell frequency. When HSC numbers are calculated, it is clear that the total number of My- HSCs increases with age and these are the predominant stem cell population present in the bone marrow. However, it is also evident that the total number of Ly-HSCs also increases with age. Why does lymphopoiesis decline with age despite the fact that Ly-HSCs do not decrease in number? This proposal will test the hypothesis that intrinsic Ly-HSC defects combined with alterations in their environment are responsible. In Aim 1, we will generate robust, whole transcriptome databases for Ly-HSCs and My-HSCs isolated from young and old mice and, using subtractive approaches, identify changes in gene expression common to both or restricted to one or the other HSC subset. These data sets will provide the rationale for subsequent experiments that determine how aging affects such key functions as proliferation, self-renewal, and DNA repair in Ly-HSCs versus My-HSCs. Chronic inflammation increases with age and has been shown to affect the proliferation, differentiation, and self-renewal of HSCs. Aim 2 will use both in vivo and in vitro models to test the hypothesis that inflammatory mediators have selective effects on Ly-HSCs and My-HSCs. Using the databases generated in Aim 1, we will identify cytokine receptors expressed on Ly-HSCs and My-HSCs and test how their ligands affect the growth and differentiation of each HSC subset using a stem cell colony assay we have developed. We will also test the hypothesis that Ly-HSCs and My-HSCs differentially process inflammatory signals due to differences in the level at which they activate NF-?B and express NF-?B regulated genes. The ability to stimulate lymphopoiesis in the elderly could have important translational implications. We have observed that treatment of old mice with Insulin-Like Growth Factor-I (IGF-I), a naturally occurring hormone whose production declines with age, can expand HSC numbers and restore the Ly-HSC/My-HSC ratio to that in young animals. Experiments in Aim 3 will assess the pattern of gene expression in Ly-HSCs harvested from the IGF-I treated mice and, using approaches defined in Aims 1 and 2, determine if the Ly-HSCs from IGF-I treated mice have been rejuvenated. Taken together, the proposed studies will provide new information about how aging impacts HSC subpopulations and pre-clinical insights into the potential of pharmacologic interventions to stimulate lymphopoiesis in the elderly.